CN101333561A - Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip - Google Patents

Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip Download PDF

Info

Publication number
CN101333561A
CN101333561A CNA200710043226XA CN200710043226A CN101333561A CN 101333561 A CN101333561 A CN 101333561A CN A200710043226X A CNA200710043226X A CN A200710043226XA CN 200710043226 A CN200710043226 A CN 200710043226A CN 101333561 A CN101333561 A CN 101333561A
Authority
CN
China
Prior art keywords
pos
gene
apob
enos
apoe
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
CNA200710043226XA
Other languages
Chinese (zh)
Inventor
穆海东
汪宁梅
杜喜玲
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Shanghai Yulong Biological Science and Technology Co Ltd
Original Assignee
Shanghai Yulong Biological Science and Technology Co Ltd
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Shanghai Yulong Biological Science and Technology Co Ltd filed Critical Shanghai Yulong Biological Science and Technology Co Ltd
Priority to CNA200710043226XA priority Critical patent/CN101333561A/en
Publication of CN101333561A publication Critical patent/CN101333561A/en
Pending legal-status Critical Current

Links

Images

Landscapes

  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention provides a cardiovascular and cerebrovascular diseases susceptibility gene chip detection kit belonging to the gene chip technique field of clinical detection, which can be used to parallelly and economically detect thirteen genes related to the susceptibility of cardiovascular and cerebrovascular diseases. The kit comprises an extraction solution, a hybridization buffer solution, a washing liquor, an amplifying solution, Taq enzyme and a gene chip, the amplifying solution contains a primer for amplifying the thirteen genes containing mutant sites, and the thirteen genes respectively are: an Enos gene, an ADD1 gene, an APOB-2515 gene, an AT1R gene, an APOB-3500 gene, an APOB-4181 gene, an eNOS-894 gene, an FgB-148 gene, an APOE gene, a GNB3 gene, an AGT gene, an FVII gene and an MTHFR gene. A plurality of genes are related to the susceptibility of the cardiovascular and cerebrovascular diseases, and the detection kit can be used for detecting the related thirteen genes parallelly.

Description

Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip
Technical field
The invention belongs to the biochip technology field of clinical detection, relate in particular to a kind of detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip.
Background technology
The tumor susceptibility gene relevant with cardiovascular and cerebrovascular diseases is many, by consulting document, picks out 13 genes that dependency is stronger, be respectively the Enos gene, ADD1 gene, APOB-2515 gene, the AT1R gene, APOB-3500 gene, APOB-4181 gene, the eNOS-894 gene, FgB-148 gene, APOE gene, the GNB3 gene, the AGT gene, FVII gene, mthfr gene.In order to cardiovascular and cerebrovascular diseases predisposing gene is carried out correlative study.
Be used for collimation to cardiovascular and cerebrovascular diseases predisposing gene research at present and do not reach 13 more than the gene, the tumor susceptibility gene of this test kit is summarized comprehensively.Detection method to the point mutation of cardiovascular and cerebrovascular diseases predisposing gene genes involved mainly contains in addition: sequencing, and this is the most direct, the most accurately method; Fluorescence quantitative PCR method and single-basic extension method.These method devices needed instruments all cost an arm and a leg, and technical sophistication is from considering that economically being unsuitable for being used as ordinary method is promoted.
Summary of the invention
For the genes involved to the cardiovascular and cerebrovascular diseases susceptible carries out parallel detection, great amount of samples is analyzed simultaneously, and the economic feasibility of promoting of consideration, the invention provides a kind of detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, comprise extract, hybridization buffer, washing lotion, amplification liquid, Taq enzyme, gene chip; It is characterized in that contained amplification liquid contains 13 genes that contain the mutational site of amplification, is respectively the Enos gene, the ADD1 gene, APOB-2515 gene, AT1R gene, the APOB-3500 gene, APOB-4181 gene, eNOS-894 gene, the FgB-148 gene, the APOE gene, GNB3 gene, AGT gene, the FVII gene, mthfr gene.
Described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, amplification liquid can comprise the primer with next group sequence::
Enos-1：CCAGATGCCCAGCTAG
Enos-2：GACATTAGGGTATCCCTT
ADD1-1：CACAGGTTACTCCTTTGCTA
ADD1-2：GGTTTCACGTGATCGTGT
APOB-2515-1：AACCAAGGCCACAGTTGC
APOB-2515-2：CTCTACCAATGCTTTCATA
AT1R-1：CATGTTCGAAACCTGTCCA
AT1R-2：TCGAGTTTCTGACATTGTTC
APOB-3500-1：AATGTCAGCTCTTGTTCAGG
APOB-3500-2：CCAGCTTTAGGAAATCC
APOB-4181-1：GGCCATTAGGCAAATTGAT
APOB-4181-2：CCTCCCTTATGAACATAGT
eNOS-894-1：ATTCAGCACGGCTGGA
eNOS-894-2：TGCAGGCCCTTCTTGA
FgB-148-1：CGGAGCTTGTGTAGTTTC
FgB-148-2：TGGAAGCTCCAAGAAACC
APOE-1：CGAGACCATGAAGGAGTT
APOE-2：CTGCCCATCTCCTCCA
GNB3-1：GATCCCTGACCCACTTG
GNB3-2：TTACCCACACGCTCAGAC
AGT-1：GTTGCTGCTGAGAAGATT
AGT-2：CCATCTCCAAGGCCTG
FVII-1：TCATGGTCCTCAACGT
FVII-2：TTTCTCTCCACAGGCC
MTHFR-1：CCTGAAGCACTTGAAGG
MTHFR-2：GGGTGGAGGGAGCTTA
Described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip can also comprise amplification liquid B, and amplification liquid B can comprise ASPE (Allele-Specific Primer Extension, the i.e. special characteristic primer extension of the allelotrope) primer as next group sequence:
Enos-T：agctcttccctggcT
Enos-C：agctcttccctggcC
ADD1-G：aagcttccgaggaaG
ADD1-T：aagcttccgaggaaT
APOB-2515-C：aaattccgagagacC
APOB-2515-T：aattccgagagacT
AT1R-A：ttcactaccaaatgagcA
AT1R-C：ttcactaccaaatgagcC
APOB-3500-G：atggaaaaatgaagtccG
APOB-3500-A：atggaaaaatgaagtccA
APOB-4181-G：ggtacgagttactcaaG
APOB-4181-A：ggtacgagttactcaaA
eNOS-894-G：aggccccagatgaG
eNOS-894-T：aggccccagatgaT
FgB-148-C：gtatgacaagtaaataagC
FgB-148-T：gtatgacaagtaaataagT
APOE--P?1：acatggaggacgtgT
APOE-P2：gacatggaggacgtgC
APOE-P3：gatgacctgcagaagC
APOE-P4：gacctgcagaagT
GNB3-C：ctgcggcatcacgtcC
GNB3-T：ctgcggcatcacgtcT
AGT-T：ctggctgctccctgaT
AGT-C：ctggctgctccctgaC
FVII-G：tgccacccactaccG
FVII-A：tgccacccactaccA
MTHFR-C：GGTGTCTGCGGGAGC
MTHFR-T：GGTGTCTGCGGGAGT
Described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, the ASPE primer that comprises among the described amplification liquid B can also be the following sequences that contains 5 '-end detection and location sequence:
Enos-tagT：CTTTAATCTCAATCAATACAAATCagctcttccctggcT
Enos-tagC：CTTTATCAATACATACTACAATCAagctcttccctggcC
ADD?1-tagG：TACACTTTATCAAATCTTACAATaagcttccgaggaaG
ADD?1-tagT：TACATTACCAATAATCTTCAAATCaagcttccgaggaaT
APOB-2515-tagC：CAATTCAAATCACAATAATCAATCaaattccgagagacC
APOB-2515-tagT：TCAACAATCTTTTACAATCAAATCaattccgagagacT
AT?1R-tagA：CAATTCATTTACCAATTTACCAATttcactaccaaatgagcA
AT?1R-tagC：AATCCTTTTACATTCATTACTTACttcactaccaaatgagcC
APOB-3500-tagG：TAATCTTCTATATCAACATCTTACatggaaaaatgaagtccG
APOB-3500-tagA：ATCATACATACATACAAATCTACAatggaaaaatgaagtccA
APOB-4181-tagG：TACAAATCATCAATCACTTTAATggtacgagttactcaaG
APOB-4181-tagA：CAATAAACTATACTTCTTCACTAAggtacgagttactcaaA
eNOS-894-tagG：CTACTATACATCTTACTATACTTTaggccccagatgaG
eNOS-894-tagT：ATACTTCATTCATTCATCAATTCAcaggccccagatgaT
FgB-148-tagC：AATCAATCTTCATTCAAATCATCAgtatgacaagtaaataagC
FgB-148-tagT：AATCCTTTCTTTAATCTCAAATCAgtatgacaagtaaataagT
APOE-tag-P?1：TTACTCAAAATCTACACTTTTTCAacatggaggacgtgT
APOE-tag-P2：CTACAAACAAACAAACATTATCAAgacatggaggacgtgC
APOE-tag-P3：TTCACTTTTCAATCAACTTTAATCgatgacctgcagaagC
APOE-tag-P4：TCAATTACTTCACTTTAATCCTTTgacctgcagaagT
GNB3-tagC：CAATTCATTTCATTCACAATCAATctgcggcatcacgtcC
GNB3-tagT：CAATATACCAATATCATCATTTACctgcggcatcacgtcT
AGT-tagT：TCAATCATCTTTATACTTCACAATctggctgctccctgaT
AGT-tagC：CTTTTTCAATCACTTTCAATTCATctggctgctccctgaC
FVII-tagG：CAATTTACTCATATACATCACTTTtgccacccactaccG
FVII-tagA：CTACTAATTCATTAACATTACTACtgccacccactaccA
MTHFR-tagC：CTTTAATCCTTTATCACTTTATCAGGTGTCTGCGGGAGC
MTHFR-tagT：TCAATCAATTACTTACTCAAATACGGTGTCTGCGGGAGT
Described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, amplification liquid B contains biotin labeled dCTP.
Described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, the probe sequence on the contained gene chip can be following sequences:
POS-1：GATTTGTATTGATTGAGATTAAAG
POS-2：TGATTGTAGTATGTATTGATAAAG
POS-3：GATTGTAAGATTTGATAAAGTGTA
POS-4：GATTTGAAGATTATTGGTAATGTA
POS-5：GATTGATTATTGTGATTTGAATTG
POS-6：GATTTGATTGTAAAAGATTGTTGA
POS-7：ATTGGTAAATTGGTAAATGAATTG
POS-8：GTAAGTAATGAATGTAAAAGGATT
POS-9：GTAAGATGTTGATATAGAAGATTA
POS-10：TGTAGATTTGTATGTATGTATGAT
POS-11：GATTAAAGTGATTGATGATTTGTA
POS-12：TTAGTGAAGAAGTATAGTTTATTG
POS-13：AAAGTATAGTAAGATGTATAGTAG
POS-14：TGAATTGATGAATGAATGAAGTAT
POS-15：TGATGATTTGAATGAAGATTGATT
POS-16：TGATAAAGTGATAAAGGATTAAAG
POS-17：GTATTTGAGTAAGTAATTGATTGA
POS-18：TGATTTGAGATTAAAGAAAGGATT
POS-19：TGAAAAAGTGTAGATTTTGAGTAA
POS-20：TTGATAATGTTTGTTTGTTTGTAG
POS-21：GATTAAAGTTGATTGAAAAGTGAA
POS-22：AAAGGATTAAAGTGAAGTAATTGA
POS-23：ATTGATTGTGAATGAAATGAATTG
POS-24：GTAAATGATGATATTGGTATATTG
POS-25：ATTGTGAAGTATAAAGATGATTGA
POS-26：ATGAATTGAAAGTGATTGAAAAAG
POS-27：AAAGTGATGTATATGAGTAAATTG
POS-28：GTAGTAATGTTAATGAATTAGTAG
The detection of 13 tumor susceptibility genes of this test kit by will be relevant with cardiovascular and cerebrovascular diseases combines, and reaches the purpose of genes involved being carried out parallel detection.By the ASPE method is combined with chemoluminescence method, detect the unit point sudden change in addition, advantage is: (1) result is accurate, and (2) flux is big, and (3) background is low, and (4) are easy and simple to handle, and (5) need not large-scale instrument.
Description of drawings
Fig. 1 is the chip inspecting reagent unit applicating flow chart
Fig. 2 is a contained cardiovascular and cerebrovascular diseases predisposing gene detection chip synoptic diagram in the test kit
Embodiment
Below in conjunction with accompanying drawing and the enforcement that illustrates the object of the invention.
Examples of implementation:
1. sample process
Originate as sample with the oral mucosa cast-off cells, carry out genome DNA extraction.The cellular genome extraction agent box of vast Tyke is used in extracting.
2. chip preparation, with existing mark position probe point sample on super flat slide in the document, probe sequence is as follows:
POS-1：GATTTGTATTGATTGAGATTAAAG
POS-2：TGATTGTAGTATGTATTGATAAAG
POS-3：GATTGTAAGATTTGATAAAGTGTA
POS-4：GATTTGAAGATTATTGGTAATGTA
POS-5：GATTGATTATTGTGATTTGAATTG
POS-6：GATTTGATTGTAAAAGATTGTTGA
POS-7：ATTGGTAAATTGGTAAATGAATTG
POS-8：GTAAGTAATGAATGTAAAAGGATT
POS-9：GTAAGATGTTGATATAGAAGATTA
POS-10：TGTAGATTTGTATGTATGTATGAT
POS-11：GATTAAAGTGATTGATGATTTGTA
POS-12：TTAGTGAAGAAGTATAGTTTATTG
POS-13：AAAGTATAGTAAGATGTATAGTAG
POS-14：TGAATTGATGAATGAATGAAGTAT
POS-15：TGATGATTTGAATGAAGATTGATT
POS-16：TGATAAAGTGATAAAGGATTAAAG
POS-17：GTATTTGAGTAAGTAATTGATTGA
POS-18：TGATTTGAGATTAAAGAAAGGATT
POS-19：TGAAAAAGTGTAGATTTTGAGTAA
POS-20：TTGATAATGTTTGTTTGTTTGTAG
POS-21：GATTAAAGTTGATTGAAAAGTGAA
POS-22：AAAGGATTAAAGTGAAGTAATTGA
POS-23：ATTGATTGTGAATGAAATGAATTG
POS-24：GTAAATGATGATATTGGTATATTG
POS-25：ATTGTGAAGTATAAAGATGATTGA
POS-26：ATGAATTGAAAGTGATTGAAAAAG
POS-27：AAAGTGATGTATATGAGTAAATTG
POS-28：GTAGTAATGTTAATGAATTAGTAG
3. design of primers:
(1) the following primer sequence group A of design is relevant with the cardiovascular and cerebrovascular diseases susceptibility in order to amplification, contains 13 gene segments in mutational site:
Enos-1：CCAGATGCCCAGCTAG
Enos-2：GACATTAGGGTATCCCTT
ADD1-1：CACAGGTTACTCCTTTGCTA
ADD1-2：GGTTTCACGTGATCGTGT
APOB-2515-1：AACCAAGGCCACAGTTGC
APOB-2515-2：CTCTACCAATGCTTTCATA
AT1R-1：CATGTTCGAAACCTGTCCA
AT1R-2：TCGAGTTTCTGACATTGTTC
APOB-3500-1：AATGTCAGCTCTTGTTCAGG
APOB-3500-2：CCAGCTTTAGGAAATCC
APOB-4181-1：GGCCATTAGGCAAATTGAT
APOB-4181-2：CCTCCCTTATGAACATAGT
eNOS-894-1：ATTCAGCACGGCTGGA
eNOS-894-2：TGCAGGCCCTTCTTGA
FgB-148-1：CGGAGCTTGTGTAGTTTC
FgB-148-2：TGGAAGCTCCAAGAAACC
APOE-1：CGAGACCATGAAGGAGTT
APOE-2：CTGCCCATCTCCTCCA
GNB3-1：GATCCCTGACCCACTTG
GNB3-2：TTACCCACACGCTCAGAC
AGT-1：GTTGCTGCTGAGAAGATT
AGT-2：CCATCTCCAAGGCCTG
FVII-1：TCATGGTCCTCAACGT
FVII-2：TTTCTCTCCACAGGCC
MTHFR-1：CCTGAAGCACTTGAAGG
MTHFR-2：GGGTGGAGGGAGCTTA
(2) the following ASPE primer sequence group B of design contains the sequence in mutational site in order to amplification, introduce simultaneously to be used for the localized flag sequence of gene test so that with chip on corresponding position probe hybridization, thereby the acquisition detected result.
Enos-tagT：CTTTAATCTCAATCAATACAAATCagctcttccctggcT
Enos-tagC：CTTTATCAATACATACTACAATCAagctcttccctggcC
ADD1-tagG：TACACTTTATCAAATCTTACAATaagcttccgaggaaG
ADD1-tagT：TACATTACCAATAATCTTCAAATCaagcttccgaggaaT
APOB-2515-tagC：CAATTCAAATCACAATAATCAATCaaattccgagagacC
APOB-2515-tagT：TCAACAATCTTTTACAATCAAATCaattccgagagacT
AT1R-tagA：CAATTCATTTACCAATTTACCAATttcactaccaaatgagcA
AT1R-tagC：AATCCTTTTACATTCATTACTTACttcactaccaaatgagcC
APOB-3500-tagG：TAATCTTCTATATCAACATCTTACatggaaaaatgaagtccG
APOB-3500-tagA：ATCATACATACATACAAATCTACAatggaaaaatgaagtccA
APOB-4181-tagG：TACAAATCATCAATCACTTTAATggtacgagttactcaaG
APOB-4181-tagA：CAATAAACTATACTTCTTCACTAAggtacgagttactcaaA
eNOS-894-tagG：CTACTATACATCTTACTATACTTTaggccccagatgaG
eNOS-894-tagT：ATACTTCATTCATTCATCAATTCAcaggccccagatgaT
FgB-148-tagC：AATCAATCTTCATTCAAATCATCAgtatgacaagtaaataagC
FgB-148-tagT：AATCCTTTCTTTAATCTCAAATCAgtatgacaagtaaataagT
APOE-tag-P?1：TTACTCAAAATCTACACTTTTTCAacatggaggacgtgT
APOE-tag-P2：CTACAAACAAACAAACATTATCAAgacatggaggacgtgC
APOE-tag-P3：TTCACTTTTCAATCAACTTTAATCgatgacctgcagaagC
APOE-tag-P4：TCAATTACTTCACTTTAATCCTTTgacctgcagaagT
GNB3-tagC：CAATTCATTTCATTCACAATCAATctgcggcatcacgtcC
GNB3-tagT：CAATATACCAATATCATCATTTACctgcggcatcacgtcT
AGT-tagT：TCAATCATCTTTATACTTCACAATctggctgctccctgaT
AGT-tagC：CTTTTTCAATCACTTTCAATTCATctggctgctccctgaC
FVII-tagG：CAATTTACTCATATACATCACTTTtgccacccactaccG
FVII-tagA：CTACTAATTCATTAACATTACTACtgccacccactaccA
MTHFR-tagC：CTTTAATCCTTTATCACTTTATCAGGTGTCTGCGGGAGC
MTHFR-tagT：TCAATCAATTACTTACTCAAATACGGTGTCTGCGGGAGT
The segment to be measured and introduce biotin labeling 4.PCR increase.Contain the sequence in mutational site to be measured with primer sequence group A amplification, contain the sequence in mutational site, add during amplification and contain biotin labeled dCTP, introduce the required biotin labeling of sample product detection with this with ASPE primer sequence group B amplification.
5. hybridization detects
According to the following steps the respective sample product that obtains is hybridized detection:
(1) (5 * SSC 0.2%SDS) with 1: 6 mixed, adds conversion zone, 50ul for marked product that amplification is obtained and hybridization solution.
(2) 42 ℃ of heat and moisture preservings reacted 60 minutes.
(3) (2 * SSC, 0.1%SDS) washing is 4-6 time, each 2 minutes, vibrates with washing lotion.
(4) the avidin reaction solution of adding HRP mark, 50ul.
(5) at 42 ℃ of reaction 15-20min.
(6) (2 * SSC, 0.1%SDS) washing is 4-6 time, each 2 minutes, rocks with washing lotion.
(7) add detection liquid 20ul/ hole, make detection liquid be uniformly distributed in conversion zone.
(8) CCD detects, and looks luminous signal intensity exposure 30-60 second.
6. the result judges:
According to the CutOff value of different indexs, reference standard product gradient curve, the yin and yang attribute of judgement sample.
Meanwhile, with ordinary method samples tested is carried out conventional sense.The result shows that the test result that draws with test kit of the present invention is identical with ordinary method.
Sequence table
Enos-1：CCAGATGCCCAGCTAG
Enos-2：GACATTAGGGTATCCCTT
ADD1-1：CACAGGTTACTCCTTTGCTA
ADD1-2：GGTTTCACGTGATCGTGT
APOB-2515-1：AACCAAGGCCACAGTTGC
APOB-2515-2：CTCTACCAATGCTTTCATA
AT1R-1：CATGTTCGAAACCTGTCCA
AT1R-2：TCGAGTTTCTGACATTGTTC
APOB-3500-1：AATGTCAGCTCTTGTTCAGG
APOB-3500-2：CCAGCTTTAGGAAATCC
APOB-4181-1：GGCCATTAGGCAAATTGAT
APOB-4181-2：CCTCCCTTATGAACATAGT
eNOS-894-1：ATTCAGCACGGCTGGA
eNOS-894-2：TGCAGGCCCTTCTTGA
FgB-148-1：CGGAGCTTGTGTAGTTTC
FgB-148-2：TGGAAGCTCCAAGAAACC
APOE-1：CGAGACCATGAAGGAGTT
APOE-2：CTGCCCATCTCCTCCA
GNB3-1：GATCCCTGACCCACTTG
GNB3-2：TTACCCACACGCTCAGAC
AGT-1：GTTGCTGCTGAGAAGATT
AGT-2：CCATCTCCAAGGCCTG
FVII-1：TCATGGTCCTCAACGT
FVII-2：TTTCTCTCCACAGGCC
MTHFR-1：CCTGAAGCACTTGAAGG
MTHFR-2：GGGTGGAGGGAGCTTA
Enos-T：agctcttccctggcT
Enos-C：agctcttccctggcC
ADD1-G：aagcttccgaggaaG
ADD1-T：aagcttccgaggaaT
APOB-2515-C：aaattccgagagacC
APOB-2515-T：aattccgagagacT
AT1R-A：ttcactaccaaatgagcA
AT1R-C：ttcactaccaaatgagcC
APOB-3500-G：atggaaaaatgaagtccG
APOB-3500-A：atggaaaaatgaagtccA
APOB-4181-G：ggtacgagttactcaaG
APOB-4181-A：ggtacgagttactcaaA
eNOS-894-G：aggccccagatgaG
eNOS-894-T：aggccccagatgaT
FgB-148-C：gtatgacaagtaaataagC
FgB-148-T：gtatgacaagtaaataagT
APOE--P1：acatggaggacgtgT
APOE-P2：gacatggaggacgtgC
APOE-P3：gatgacctgcagaagC
APOE-P4：gacctgcagaagT
GNB3-C：ctgcggcatcacgtcC
GNB3-T：ctgcggcatcacgtcT
AGT-T：ctggctgctccctgaT
AGT-C：ctggctgctccctgaC
FVII-G：tgccacccactaccG
FVII-A：tgccacccactaccA
MTHFR-C：GGTGTCTGCGGGAGC
MTHFR-T：GGTGTCTGCGGGAGT
Enos-tagT：CTTTAATCTCAATCAATACAAATCagctcttccctggcT
Enos-tagC：CTTTATCAATACATACTACAATCAagctcttccctggcC
ADD1-tagG：TACACTTTATCAAATCTTACAATaagcttccgaggaaG
ADD1-tagT：TACATTACCAATAATCTTCAAATCaagcttccgaggaaT
APOB-2515-tagC：CAATTCAAATCACAATAATCAATCaaattccgagagacC
APOB-2515-tagT：TCAACAATCTTTTACAATCAAATCaattccgagagacT
AT1R-tagA：CAATTCATTTACCAATTTACCAATttcactaccaaatgagcA
AT1R-tagC：AATCCTTTTACATTCATTACTTACttcactaccaaatgagcC
APOB-3500-tagG：TAATCTTCTATATCAACATCTTACatggaaaaatgaagtccG
APOB-3500-tagA：ATCATACATACATACAAATCTACAatggaaaaatgaagtccA
APOB-4181-tagG：TACAAATCATCAATCACTTTAATggtacgagttactcaaG
APOB-4181-tagA：CAATAAACTATACTTCTTCACTAAggtacgagttactcaaA
eNOS-894-tagG：CTACTATACATCTTACTATACTTTaggccccagatgaG
eNOS-894-tagT：ATACTTCATTCATTCATCAATTCAcaggccccagatgaT
FgB-148-tagC：AATCAATCTTCATTCAAATCATCAgtatgacaagtaaataagC
FgB-148-tagT：AATCCTTTCTTTAATCTCAAATCAgtatgacaagtaaataagT
APOE-tag-P1：TTACTCAAAATCTACACTTTTTCAacatggaggacgtgT
APOE-tag-P2：CTACAAACAAACAAACATTATCAAgacatggaggacgtgC
APOE-tag-P3：TTCACTTTTCAATCAACTTTAATCgatgacctgcagaagC
APOE-tag-P4：TCAATTACTTCACTTTAATCCTTTgacctgcagaagT
GNB3-tagC：CAATTCATTTCATTCACAATCAATctgcggcatcacgtcC
GNB3-tagT：CAATATACCAATATCATCATTTACctgcggcatcacgtcT
AGT-tagT：TCAATCATCTTTATACTTCACAATctggctgctccctgaT
AGT-tagC：CTTTTTCAATCACTTTCAATTCATctggctgctccctgaC
FVII-tagG：CAATTTACTCATATACATCACTTTtgccacccactaccG
FVII-tagA：CTACTAATTCATTAACATTACTACtgccacccactaccA
MTHFR-tagC：CTTTAATCCTTTATCACTTTATCAGGTGTCTGCGGGAGC
MTHFR-tagT：TCAATCAATTACTTACTCAAATACGGTGTCTGCGGGAGT
POS-1：GATTTGTATTGATTGAGATTAAAG
POS-2：TGATTGTAGTATGTATTGATAAAG
POS-3：GATTGTAAGATTTGATAAAGTGTA
POS-4：GATTTGAAGATTATTGGTAATGTA
POS-5：GATTGATTATTGTGATTTGAATTG
POS-6：GATTTGATTGTAAAAGATTGTTGA
POS-7：ATTGGTAAATTGGTAAATGAATTG
POS-8：GTAAGTAATGAATGTAAAAGGATT
POS-9：GTAAGATGTTGATATAGAAGATTA
POS-10：TGTAGATTTGTATGTATGTATGAT
POS-11：GATTAAAGTGATTGATGATTTGTA
POS-12：TTAGTGAAGAAGTATAGTTTATTG
POS-13：AAAGTATAGTAAGATGTATAGTAG
POS-14：TGAATTGATGAATGAATGAAGTAT
POS-15：TGATGATTTGAATGAAGATTGATT
POS-16：TGATAAAGTGATAAAGGATTAAAG
POS-17：GTATTTGAGTAAGTAATTGATTGA
POS-18：TGATTTGAGATTAAAGAAAGGATT
POS-19：TGAAAAAGTGTAGATTTTGAGTAA
POS-20：TTGATAATGTTTGTTTGTTTGTAG
POS-21：GATTAAAGTTGATTGAAAAGTGAA
POS-22：AAAGGATTAAAGTGAAGTAATTGA
POS-23：ATTGATTGTGAATGAAATGAATTG
POS-24：GTAAATGATGATATTGGTATATTG
POS-25：ATTGTGAAGTATAAAGATGATTGA
POS-26：ATGAATTGAAAGTGATTGAAAAAG
POS-27：AAAGTGATGTATATGAGTAAATTG
POS-28：GTAGTAATGTTAATGAATTAGTAG

Claims (6)

1. a detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip comprises extract, hybridization buffer, washing lotion, amplification liquid, Taq enzyme, gene chip; It is characterized in that contained amplification liquid contains 13 genes that contain the mutational site of amplification, is respectively the Enos gene, the ADD1 gene, APOB-2515 gene, AT1R gene, the APOB-3500 gene, APOB-4181 gene, eNOS-894 gene, the FgB-148 gene, the APOE gene, GNB3 gene, AGT gene, the FVII gene, mthfr gene.
2. detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip according to claim 1 is characterized in that, described amplification liquid can comprise the primer with next group sequence:
Enos-1：CCAGATGCCCAGCTAG
Enos-2：GACATTAGGGTATCCCTT
ADD1-1：CACAGGTTACTCCTTTGCTA
ADD1-2：GGTTTCACGTGATCGTGT
APOB-2515-1：AACCAAGGCCACAGTTGC
APOB-2515-2：CTCTACCAATGCTTTCATA
AT1R-1：CATGTTCGAAACCTGTCCA
AT1R-2：TCGAGTTTCTGACATTGTTC
APOB-3500-1：AATGTCAGCTCTTGTTCAGG
APOB-3500-2：CCAGCTTTAGGAAATCC
APOB-4181-1：GGCCATTAGGCAAATTGAT
APOB-4181-2：CCTCCCTTATGAACATAGT
eNOS-894-1：ATTCAGCACGGCTGGA
eNOS-894-2：TGCAGGCCCTTCTTGA
FgB-148-1：CGGAGCTTGTGTAGTTTC
FgB-148-2：TGGAAGCTCCAAGAAACC
APOE-1：CGAGACCATGAAGGAGTT
APOE-2：CTGCCCATCTCCTCCA
GNB3-1：GATCCCTGACCCACTTG
GNB3-2：TTACCCACACGCTCAGAC
AGT-1：GTTGCTGCTGAGAAGATT
AGT-2：CCATCTCCAAGGCCTG
FVII-1：TCATGGTCCTCAACGT
FVII-2：TTTCTCTCCACAGGCC
MTHFR-1：CCTGAAGCACTTGAAGG
MTHFR-2：GGGTGGAGGGAGCTTA
3. detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip according to claim 1 and 2, it is characterized in that, can also comprise amplification liquid B, amplification liquid B can comprise ASPE (Allele-Specific Primer Extension, the i.e. special characteristic primer extension of the allelotrope) primer as next group sequence:
Enos-T：agctcttccctggcT
Enos-C：agctcttccctggcC
ADD1-G：aagcttccgaggaaG
ADD1-T：aagcttccgaggaaT
APOB-2515-C：aaattccgagagacC
APOB-2515-T：aattccgagagacT
AT1R-A：ttcactaccaaatgagcA
AT1R-C：ttcactaccaaatgagcC
APOB-3500-G：atggaaaaatgaagtccG
APOB-3500-A：atggaaaaatgaagtccA
APOB-4181-G：ggtacgagttactcaaG
APOB-4181-A：ggtacgagttactcaaA
eNOS-894-G：aggccccagatgaG
eNOS-894-T：aggccccagatgaT
FgB-148-C：gtatgacaagtaaataagC
FgB-148-T：gtatgacaagtaaataagT
APOE--P1：acatggaggacgtgT
APOE-P2：gacatggaggacgtgC
APOE-P3：gatgacctgcagaagC
APOE-P4：gacctgcagaagT
GNB3-C：ctgcggcatcacgtcC
GNB3-T：ctgcggcatcacgtcT
AGT-T：ctggctgctccctgaT
AGT-C：ctggctgctccctgaC
FVII-G：tgccacccactaccG
FVII-A：tgccacccactaccA
MTHFR-C：GGTGTCTGCGGGAGC
MTHFR-T：GGTGTCTGCGGGAGT
4. detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip according to claim 3 is characterized in that, the ASPE primer that comprises among the described amplification liquid B can also be the following sequence that contains 5 '-end detection and location sequence:
Enos-tagT：CTTTAATCTCAATCAATACAAATCagctcttccctggcT
Enos-tagC：CTTTATCAATACATACTACAATCAagctcttccctggcC
ADD1-tagG：TACACTTTATCAAATCTTACAATaagcttccgaggaaG
ADD1-tagT：TACATTACCAATAATCTTCAAATCaagcttccgaggaaT
APOB-2515-tagC：CAATTCAAATCACAATAATCAATCaaattccgagagacC
APOB-2515-tagT：TCAACAATCTTTTACAATCAAATCaattccgagagacT
AT1R-tagA：CAATTCATTTACCAATTTACCAATttcactaccaaatgagcA
AT1R-tagC：AATCCTTTTACATTCATTACTTACttcactaccaaatgagcC
APOB-3500-tagG：TAATCTTCTATATCAACATCTTACatggaaaaatgaagtccG
APOB-3500-tagA：ATCATACATACATACAAATCTACAatggaaaaatgaagtccA
APOB-4181-tagG：TACAAATCATCAATCACTTTAATggtacgagttactcaaG
APOB-4181-tagA：CAATAAACTATACTTCTTCACTAAggtacgagttactcaaA
eNOS-894-tagG：CTACTATACATCTTACTATACTTTaggccccagatgaG
eNOS-894-tagT：ATACTTCATTCATTCATCAATTCAcaggccccagatgaT
FgB-148-tagC：AATCAATCTTCATTCAAATCATCAgtatgacaagtaaataagC
FgB-148-tagT：AATCCTTTCTTTAATCTCAAATCAgtatgacaagtaaataagT
APOE-tag-P1：TTACTCAAAATCTACACTTTTTCAacatggaggacgtgT
APOE-tag-P2：CTACAAACAAACAAACATTATCAAgacatggaggacgtgC
APOE-tag-P3：TTCACTTTTCAATCAACTTTAATCgatgacctgcagaagC
APOE-tag-P4：TCAATTACTTCACTTTAATCCTTTgacctgcagaagT
GNB3-tagC：CAATTCATTTCATTCACAATCAATctgcggcatcacgtcC
GNB3-tagT：CAATATACCAATATCATCATTTACctgcggcatcacgtcT
AGT-tagT：TCAATCATCTTTATACTTCACAATctggctgctccctgaT
AGT-tagC：CTTTTTCAATCACTTTCAATTCATctggctgctccctgaC
FVII-tagG：CAATTTACTCATATACATCACTTTtgccacccactaccG
FVII-tagA：CTACTAATTCATTAACATTACTACtgccacccactaccA
MTHFR-tagC：CTTTAATCCTTTATCACTTTATCAGGTGTCTGCGGGAGC
MTHFR-tagT：TCAATCAATTACTTACTCAAATACGGTGTCTGCGGGAGT
5. according to claim 3 or 4 described detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip, it is characterized in that described amplification liquid B contains biotin labeled dCTP.
6. detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip according to claim 4 is characterized in that, the probe sequence on the contained gene chip can be following sequence:
POS-1：GATTTGTATTGATTGAGATTAAAG
POS-2：TGATTGTAGTATGTATTGATAAAG
POS-3：GATTGTAAGATTTGATAAAGTGTA
POS-4：GATTTGAAGATTATTGGTAATGTA
POS-5：GATTGATTATTGTGATTTGAATTG
POS-6：GATTTGATTGTAAAAGATTGTTGA
POS-7：ATTGGTAAATTGGTAAATGAATTG
POS-8：GTAAGTAATGAATGTAAAAGGATT
POS-9：GTAAGATGTTGATATAGAAGATTA
POS-10：TGTAGATTTGTATGTATGTATGAT
POS-11：GATTAAAGTGATTGATGATTTGTA
POS-12：TTAGTGAAGAAGTATAGTTTATTG
POS-13：AAAGTATAGTAAGATGTATAGTAG
POS-14：TGAATTGATGAATGAATGAAGTAT
POS-15：TGATGATTTGAATGAAGATTGATT
POS-16：TGATAAAGTGATAAAGGATTAAAG
POS-17：GTATTTGAGTAAGTAATTGATTGA
POS-18：TGATTTGAGATTAAAGAAAGGATT
POS-19：TGAAAAAGTGTAGATTTTGAGTAA
POS-20：TTGATAATGTTTGTTTGTTTGTAG
POS-21：GATTAAAGTTGATTGAAAAGTGAA
POS-22：AAAGGATTAAAGTGAAGTAATTGA
POS-23：ATTGATTGTGAATGAAATGAATTG
POS-24：GTAAATGATGATATTGGTATATTG
POS-25：ATTGTGAAGTATAAAGATGATTGA
POS-26：ATGAATTGAAAGTGATTGAAAAAG
POS-27：AAAGTGATGTATATGAGTAAATTG
POS-28：GTAGTAATGTTAATGAATTAGTAG。
CNA200710043226XA 2007-06-29 2007-06-29 Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip Pending CN101333561A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CNA200710043226XA CN101333561A (en) 2007-06-29 2007-06-29 Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CNA200710043226XA CN101333561A (en) 2007-06-29 2007-06-29 Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip

Publications (1)

Publication Number Publication Date
CN101333561A true CN101333561A (en) 2008-12-31

Family

ID=40196441

Family Applications (1)

Application Number Title Priority Date Filing Date
CNA200710043226XA Pending CN101333561A (en) 2007-06-29 2007-06-29 Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip

Country Status (1)

Country Link
CN (1) CN101333561A (en)

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101565749B (en) * 2009-04-15 2012-01-11 广州益善生物技术有限公司 CYP2C19 and ABCB1 gene SNP detection liquid-phase chip and detection method thereof
WO2012089148A1 (en) * 2010-12-31 2012-07-05 深圳华大基因科技有限公司 Method for single cell genome analysis and kit therefor
CN102827941A (en) * 2012-09-06 2012-12-19 温州医学院附属第一医院 Gene chip testing kit for recurrent nasopharyngeal carcinoma
CN106939331A (en) * 2016-01-05 2017-07-11 中国人民解放军空军总医院 One group is used to detect pilot's LOD tumor susceptibility genes pleomorphism site and detection kit

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101565749B (en) * 2009-04-15 2012-01-11 广州益善生物技术有限公司 CYP2C19 and ABCB1 gene SNP detection liquid-phase chip and detection method thereof
WO2012089148A1 (en) * 2010-12-31 2012-07-05 深圳华大基因科技有限公司 Method for single cell genome analysis and kit therefor
US9238840B2 (en) 2010-12-31 2016-01-19 BGI-Shenzhen Co., Limited Method for single cell genome analysis and kit therefor
CN102827941A (en) * 2012-09-06 2012-12-19 温州医学院附属第一医院 Gene chip testing kit for recurrent nasopharyngeal carcinoma
CN106939331A (en) * 2016-01-05 2017-07-11 中国人民解放军空军总医院 One group is used to detect pilot's LOD tumor susceptibility genes pleomorphism site and detection kit
CN106939331B (en) * 2016-01-05 2019-08-20 中国人民解放军空军特色医学中心 One group for detecting pilot's LOD tumor susceptibility gene polymorphic site and detection kit

Similar Documents

Publication Publication Date Title
CN103045591B (en) HLA gene specific PCR amplification primer, HLA typing method and kit
CN101824466B (en) Specific primer, liquid-phase chip and method for SNP detection of CYP2C9 and VKORC1 genes
CN101812511B (en) CYP3A4 gene SNP detection specific primer, liquid-phase chip and method
JP2006518194A5 (en)
CN107227361A (en) Primer, probe and detection kit for detecting CYP2C19 gene pleiomorphisms
CN102234685B (en) Liquid phase chip for detecting PIK3CA (phosphoinositide-3-kinase, catalytic, alpha) gene mutation
CN107217107A (en) The method and kit of a kind of quick detection CYP2C19 gene pleiomorphisms
CN101892317A (en) HLA high-resolution gene sequencing kit
CN105039322B (en) DNA sequence labels and sequencing library construction method and kit
CN106498036A (en) A kind of construction method in the Drug Discovery SNP variations library for high-flux sequence detection and its application
CN102146479A (en) Primers and probes for detecting genes associated with schizophrenia, bipolar affective disorders and major depression and kits and preparation methods thereof
RU2012126088A (en) DETECTION OF AAD-12-EVENT 416 IN SOY
CN113151440A (en) Kit for predicting aspirin curative effect and adverse reaction, detection method and application thereof
CN101333561A (en) Detecting kit for cardiovascular and cerebrovascular diseases predisposing gene chip
CN107447000A (en) A kind of SNP marker related to the more lambs of sheep and its application
CN102399900B (en) Gene polymorphism detection method and kit
CN103045717B (en) Pyrosequencing techniques detects the method for mycobacterium tuberculosis detection of rifampin resistant
CN102140509B (en) Gene mutation detection method based on nucleic acid amplification on solid carrier
JP6164683B2 (en) Method for analyzing deletion site of spermatogenesis region on Y chromosome and Y chromosome
CN103045716B (en) Method for detecting isoniazid drug resistance of mycobacterium tuberculosis by using pyrosequencing technology
CN110564834B (en) Marker for distinguishing gingival stem cells and deciduous tooth pulp stem cells and application thereof
CN111560442A (en) Compound amplification kit for paternity test and individual identification of cattle and application
CN101548010A (en) Method of determining the haplotype of multiple allelic genes
CN102212621A (en) Detection kit and detection method for hepatitis B virus genotyping
CN103757094A (en) Fluorescent PCR based method for detecting polymorphism of TYMS gene

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C02 Deemed withdrawal of patent application after publication (patent law 2001)
WD01 Invention patent application deemed withdrawn after publication

Open date: 20081231